The mammalian nervous system does not regenerate after injury despite the fact that there are many molecules present which encourage/promote axonal (nerve) growth. It is believed that the lack of regeneration caused by the presence of molecules in the central nervous system (CNS) and the peripheral nervous system (PNS) which actively prevent/inhibit regeneration. Hence, the well documented inability of the adult mammalian CNS to regenerate after injury is believed to result from a predominance of inhibitory molecules.
It has been demonstrated that when neurons are grown on tissue sections of the CNS they fail to extend processes onto areas of white matter, myelin. It is believed that myelin-specific inhibitory molecules can largely account for the lack of CNS regeneration and their identification will help in the design of therapies to encourage regrowth after injury. The precise molecules responsible for this inhibition have, so far, remained elusive. If these inhibitory molecules can be identified and blocked, then neural regeneration can be encouraged.
Schwab and co-workers have identified two components in CNS myelin, in the molecular weight ranges of approximately 35 kD and 250 kD, which arrest axonal growth. The most compelling observation in support of the inhibitory action of these two protein fractions is that antibodies raised to proteins eluted from these regions of polyacrylamide gels after separation of CNS myelin proteins, specifically reverses the inhibitory effect of myelin in vitro and allows limited spinal cord regeneration when applied in vivo to transected nerves (Caroni, P. and Schwab, M. E., Neuron, 1, pp. 85-96 (1988a); J. Cell Biol., 106, pp. 1281-88 (1988b); Schnell, L. and Schwab, M. E., Nature, 343, pp. 269-72 (1990)). The nature of these two proteins and how they act have not yet been described, but, it is generally accepted that they are significant contributors to the inhibitory effect of this tissue. However, as acknowledged by the authors, other factors are likely to contribute to the inhibition by CNS myelin as even in the presence of antibodies directed against these two proteins, the majority of axons in vivo fail to regenerate (Schnell, L. and Schwab, M. E., Nature, 343, pp. 269-72 (1990); Schnell et al., Nature, 367, pp. 170-73 (1993)).
In addition to inhibitory molecules in myelin, another family of proteins has recently been identified whose members inhibit axonal regeneration. These molecules are called collapsins (Luo et al., Cell, 75, pp. 217-27 (1993)). However, collapsins are found ubiquitously throughout the nervous system and as they are found in regions of the nervous system in which axons will grow, i.e. gray matter, they are unlikely to contribute significantly to the lack of neural regeneration after injury. Instead, the collapsins most likely play a role in guiding growing axons during development.
Previously it was shown that MAG, like many members of the Ig-superfamily of molecules, could promote neurite outgrowth, in this case, from dorsal root ganglion (DRG) neurons from 2 day old rats (Johnson et al., Neuron, 3, pp. 377-85 (1989)). We observed a similar effect on DRG neurons from rats up to postnatal day 3, but after this age MAG had the opposite effect, i.e., it inhibited neurite outgrowth (Mukhopadhyay et al., Neuron, 13, pp. 757-67 (1994)). Furthermore, we also found that MAG dramatically inhibited neurite outgrowth from cerebellar neurons from rats of all ages up to adult. Polyclonal antibodies directed against MAG could specifically block both stimulatory and inhibitory effects of MAG on neurite outgrowth. MAG, therefore, depending on the age and the type of neuron, can either promote or inhibit neurite outgrowth. Subsequent to our report on the inhibitory effects of MAG, another group demonstrated, using a different complementary approach, that MAG is an inhibitor of axonal growth (McKerracher et al., Neuron, 13, pp. 805-811 (1994); WO 95/22344 (Aug. 24, 1995); incorporated herein by reference).
It would be useful to block the inhibitors of axonal regeneration for treating patients with nervous system injuries where neural regeneration is a problem. No molecule had been identified in myelin which is a potent inhibitor of axonal regeneration. Although Schwab and co-workers identified components in myelin that are inhibitory, the precise nature of these components has not been identified, i.e., they have not been cloned nor have the proteins been purified. In addition, there was no information available on the component on the neuron that the putative inhibitory molecules interact with to prevent regrowth. As no inhibitory nor interacting molecules had been precisely identified, it was difficult, if not impossible, to logically design strategies whereby these molecules can been blocked and prevented from inhibiting neural regeneration.